Glial cells are the predominant cell type in the brain, constituting more than 50% of the total cell count and outnumbering neurons 10-fold (Schubert, 1984, Developmental Biology of Cultured Nerve, Muscle, and Glia. Wiley and Sons, New York; Rutka et al., 1997, J. Neurosurg. 87, 420-430). Two major subtypes of glia are distinguished by their distinct functions in the nervous system: oligodendrocytes, which form myelin sheaths around nerve cell axons, and astrocytes, which maintain brain homeostasis and respond to pathogens and brain injury (Benveniste, 1992, Am. J. Physiol. 263, C1-16; Verkhratsky et al., 1998, Physiol. Rev. 78, 100-130).
Historically, astrocytes were considered to provide mostly passive support to neurons and the overall function of the nervous system. Recent studies question this hypothesis and suggest that astrocytes also play a critical role in several aspects of signal transmission and that defects in these functions may lead to neurodegeneration (Choi, 1988; Neuron 1, 623-634; Rothstein et al., 1996, Neuron 16, 675-686; Verkhratsky et al., 1998, Physiol. Rev. 78, 100-130; Anderson & Swanson, 2000, Glia 32, 1-14). For example, the high-affinity excitatory amino acid transport systems acting through astrocyte-specific transporters EAAT1 and EAAT2 are thought to be primarily responsible for maintenance of low levels of free intrasynaptic L-glutamate, the major excitatory neurotransmitter in the brain (Choi, 1988; Neuron 1, 623-634; Benveniste, 1992, Am. J. Physiol. 263, C1-16; Anderson & Swanson, 2000, Glia 32, 1-14). Defects that specifically abrogate this function result in accumulation of extracellular glutamate in synaptic clefts and overexcitation and death of neurons, a phenomenon referred to as excitotoxicity (Choi, 1988; Neuron 1, 623-634; Gegelashvili, G. & Schousboe, A. (1997). Molec. Pharmacol., 52, 6-15; Tanaka et al., 1997, Science 276, 1699-1702).
At least one neurodegenerative disease, Amyotropic Lateral Sclerosis (ALS), has been linked to a significant decrease of high-affinity sodium-dependent glutamate transport in synaptic membranes (Rothstein et al., 1992, New Engl. J. Med. 326, 1464-1468) and a selective loss of the transporter EAAT2 (Bristol & Rothstein, 1996, Ann. Neurol. 39, 676-679). Similar defects are implicated in Alzheimer's disease, stroke/ischemia, epilepsy, and HIV-1-associated dementia (HAD) (Choi, 1988, Neuron 1, 623-634; Kaul et al., 2001, Nature 410, 988-994; Maragakis & Rothstein, 2001, Arch. Neurol., 58, 365-370).
In addition to their role in glutamate removal from synapses, astrocytes significantly increase the number of synapses and enhance synaptic efficacy by altering pre- and post-synaptic functions in vitro (Oliet et al., 2001, Science 292, 923-926; Ullian et al., 2001, Science 291, 657-660). Finally, astrocytes appear to share many properties with neurons, including expression of functional neuronal nicotinic acetylcholine receptors (nACHRs) and competence for Ca++-dependent glutamate release, thus permitting intercellular signaling between astrocytes and neurons and, possibly, modulation of neuronal signal transmission by astrocytes (Iino et al., 2001, Science 292, 926-929; Sharma & Vijayaraghavan, 2001, Proc Natl Acad Sci USA 98, 4148-4153; Ullian et al., 2001, Science 291, 657-660).
Overall, these studies suggest that astrocytes and neurons are functionally integrated and that pathogenic stimuli that adversely affect astrocytes will directly or indirectly impact on neuronal function and survival.
Studies have focused on investigation of HIV-1 infection in neural cells and the potential contribution of such infections to neurodegeneration and HAD. The pathogenic events triggered by HIV-1 in the brain, which ultimately result in neuronal loss and CNS dysfunction (Navia et al., 1986a, Ann. Neurol. 19, 525-535; Navia et al., 1986, Ann. Neurol. 19, 517-524; reviewed in Lipton & Gendelman, 1995, New Engl. J. Med. 233, 934-940), have not been fully resolved. Neurons are rarely infected in vivo (Wiley et al., 1986, Proc. Natl. Acad. Sci. USA 83, 7089-93) and it is unlikely that neuronal loss in HIV-1 dementia is caused by cytopathic infection of these cells.
Numerous neuropathology, immunocytochemistry, in situ hybridization, and virus isolation studies indicate that macrophages and microglial cells are the primary host cells for productive HIV-1 infection in the CNS (Koening et al., 1986, Science 233, 1089-1093; Brew et al., 1995, Ann. Neurol. 38, 563-570; reviewed in Lipton & Gendelman, 1995, New Engl. J. Med. 233, 934-940). It has been suggested that HV-1 infection and subsequent activation of infected cells causes neuroinflammatory responses involving production of chemokines, cytokines, nitric oxide, and other factors, some of which were shown to be neurotoxic in vitro (reviewed in Lipton & Gendelman, 1995, New Engl. J. Med. 233, 934-940; Kaul et al., 2001, Nature 410, 988-994). Viral products secreted by infected cells, including gp120 and Tat, can also induce neurotoxicity in vitro and in animal models (Lipton & Gendelman, 1995, New Engl. J. Med. 233, 934-940; Kaul et al., 2001, Nature 410, 988-994).
Astrocytes also can be infected with HIV-1 in vitro and in vivo, although with lower efficiency than T cells and macrophages (Dewhurst et al., 1987, J. Virol. 61, 3774-3782, J. Virol. 61, 3774-3782; Tomatore et al., 1991, J. Virol. 65, 6094-6100; Saito et al., 1994, Neurology 44, 474-481; Tomatore et al., 1994, Neurology 44, 481-487; reviewed in Brack-Wemer, 1999, AIDS 13, 1-22). The limited infection of astrocytes has been attributed to various mechanisms including intracellular restrictions to virus expression (Tomatore et al., 1994b, J. Virol. 68, 93-102; Gorry et al., 1999, J. Virol. 73, 352-61; Ludwig et al., 1999, J. Virol. 73, 8279-8289) or, as has been shown recently, inefficient virus entry (Bencheikh et al., 1999, J. Neurovirol. 5, 115-124; Canki et al., 2001, J. Virol. 75, 7925-7933). There is general agreement, however, that HIV-1 can persist in astrocytes for prolonged periods in a low productive, non-cytolytic state, from which it can be induced by physiologic stimuli such as tumor necrosis factor-α (TNF-α) (Tornatore et al., 1991, J. Virol. 65, 6094-6100; Shahabuddin et al., 1992, Pathobiology 60, 195-205).
Surveys of autopsy tissues using in situ PCR and sensitive immunocytochemistry techniques indicate that the frequency of HIV-1-positive astrocytes in selected tissue sections from brains of patients with dementia can achieve 1% (Saito et al., 1994, Neurology 44, 474-481; Tomatore et al., 1994a, Neurology 44, 481-487; Takahashi et al., 1996, Ann. Neurol. 39, 705-711). Considering that the number of astrocytes in the brain is between 1011 to 1012 cells (Verkhratsky et al., 1998, Physiol. Rev. 78, 100-130), these cells clearly constitute a major target for HIV-1 infection in the brain.
The consequences of this infection with respect to HAD pathogenesis are unknown but they may be significant. Persistent, non-cytolytic HIV-1 infection in culture alters gene expression in lymphocytes (Shahabuddin et al., 1994, AIDS Res. Hum. Retroviruses 10, 1525-1529; Geiss et al., 2000, Virology 266, 8-16) and astrocytes (Schneider-Schaulies et al., 1992, Virology 191, 765-772; He et al., 1997, Proc. Natl. Acad. Sci. USA 94, 3954-3959), indicating that such infections may affect cell function. Exposure of astrocytes to recombinant HIV-1 envelope glycoprotein gp120 alters cell physiology (Benos et al., 1994a, Adv. Neuroimmunol. 4, 175-179), including a potential effect on glutamate transport as indicated by increased D-aspartate efflux in astrocytes treated with gp120 (Benos et al., 1994b, Proc. Natl. Acad. Sci. USA 91, 494-498). Impairment of glutamate transport was also observed after incubation of human astrocytes with TNF-α (Fine et al., 1996, J. Biol. Chem. 271, 15303-15306) or co-cultivation with T cells infected with human T cell leukemia virus type I (HTLVI) (Szymocha et al., 2000, J. Virol. 74, 6433-6441), and similar defects were found in feline astrocytes after infection with feline immunodeficiency virus (FIV) (Yu et al., 1998, Proc. Natl. Acad. Sci. USA 95, 2624-2629).
More recent studies indicate that ligation of the HIV-1 coreceptor on astrocytes, CXCR4, by either stromal cell-derived factor 1 (SDF-1) or gp120 can stimulate a novel signaling pathway that involves Ca2+-dependent release of glutamate (Sharma & Vijayaraghavan, 2001, Proc Natl Acad Sci USA 98, 4148-4153) in a process including activation of the CXCR4 receptor, an autocrine/paracrine TNF-α-dependent signaling, and prostaglandin (Bezzi et al., 2001, Nat. Neurosci. 4, 702-710). These results suggest that HIV-1, gp 120, and other neuropathogenic agents can alter specific signaling pathways in astrocytes in a way that may impair important physiological functions of these cells in neuronal signal transmission and response to brain injury.